Dental caries is considered to be a major health concern in the United States. Accordingly, developing drugs to prevent and control dental caries is now the `mantra' in American Dentistry. Fluoride has been used in community water supplies and toothpastes to control dental caries over the years. Though these preventive measures are successful to some extent, it does not prevent caries formation completely. Thus, new innovative approaches are necessary to combat dental caries. Although a large number of oral bacteria have been associated with dental caries, Streptococcus mutans has been implicated as the major etiological agent in the initiation and development of dental caries. One of S. mutans virulence factors attributed to its cariogenic property is its expression of extracellular glucosyl transferases (GTFs). Three distinct gtf genes expressing GTF activity are found in most strains of S. mutans. Genes gtfB and gtfC express GTF-I and GTF-SI enzymes respectively, which synthesize predominantly water- insoluble glucans and gtfD, encodes for GTF-S, an enzyme that synthesizes water-soluble glucans. Glucans contribute to the structural integrity of biofilms and provide firm adherence of growing bacteria to tooth surfaces. S. mutans defective in either or both of genes (gtfB- and gtfC-) exhibited markedly reduced levels of smooth surface caries lesion as compared to the parental S. mutans. In our own studies, we have shown that the gtfB- knockout S. mutans exhibited 80% reduction in its ability to form biofilm as compared to the wild type. Thus, functions of S. mutans GTFs are essential for glucan synthesis, biofilm formation and resulting cariogenesis. Therefore, we propose inhibition of S. mutans GTFs as a novel strategy to prevent the glucan formation and cariogenic biofilms. Unlike traditional antibiotics, S mutans GTF inhibitors will selectively inhibit cariogenic biofilms without affecting commensal oral bacterial flora. We have utilized the recently published X-ray crystal structure of GTF-SI for the design of GTF inhibitors. With the help of in-silico screening and SAR studies, we have identified low micromolar inhibitors of GTFs of S. mutans. These compounds selectively inhibit S. mutans biofilms at low micromolar concentrations without affecting the growth of S. mutans or other oral commensal bacteria. Structurally similar analogs of our lead compounds that were inactive against GTFs were also found to be inactive against biofilm, indicating that these effects are directly related. The goal of this proposal is to optimize the anti-biofilm activity of the lea compounds and to establish their potential as agents to treat cariogenic biofilms and dental caries. The specific aims are: Aim 1: To optimize the biofilm inhibitory activity of the lead compounds by structure activity relationship studies. Aim 2: To evaluate the in vitro activities of the compounds and to determine the efficacy of 1 or 2 optimized lead compounds in a rat model of dental caries.